U.S. patent number 7,070,680 [Application Number 10/514,236] was granted by the patent office on 2006-07-04 for biosensor.
This patent grant is currently assigned to Infopia Co., Ltd.. Invention is credited to Byung-Woo Bae, Mi-Joong Kwon, Heon-Kwon Lee, Sung-Dong Lee.
United States Patent |
7,070,680 |
Bae , et al. |
July 4, 2006 |
Biosensor
Abstract
A biosensor for quantifying a specific substance contained in a
biological sample includes an electrically insulating base plate, a
plurality of lead terminals formed on the base plate, a plurality
of lead wires connected to the lead terminals, respectively, an
electrode system including two working electrodes and one reference
electrode connected to the lead wires, respectively, an insulating
layer that insulates the electrodes, an enzyme reaction layer
formed on the insulating layer and the electrodes, a spacer formed
on the enzyme reaction layer so as to ensure a sufficient space
that receives a sample, and a cover formed on the spacer. Further,
the spacer has a sample introduction port opened at one side of the
spacer, the cover has at least one slit for venting air existing in
a sample receiving space defined by the spacer and the cover, and
the slit extends to above the electrodes from one end of the
cover.
Inventors: |
Bae; Byung-Woo (Kyunggi-Do,
KR), Lee; Heon-Kwon (Kyunggi-Do, KR), Lee;
Sung-Dong (Seoul, KR), Kwon; Mi-Joong (Seoul,
KR) |
Assignee: |
Infopia Co., Ltd. (Anyang-si,
KR)
|
Family
ID: |
29546312 |
Appl.
No.: |
10/514,236 |
Filed: |
October 4, 2002 |
PCT
Filed: |
October 04, 2002 |
PCT No.: |
PCT/KR02/01854 |
371(c)(1),(2),(4) Date: |
November 19, 2004 |
PCT
Pub. No.: |
WO03/097866 |
PCT
Pub. Date: |
November 27, 2003 |
Prior Publication Data
|
|
|
|
Document
Identifier |
Publication Date |
|
US 20050161323 A1 |
Jul 28, 2005 |
|
Foreign Application Priority Data
|
|
|
|
|
May 20, 2002 [KR] |
|
|
10-2002-0027971 |
|
Current U.S.
Class: |
204/403.04;
204/403.14 |
Current CPC
Class: |
C12Q
1/001 (20130101); G01N 27/3272 (20130101) |
Current International
Class: |
G01N
27/327 (20060101) |
Field of
Search: |
;204/403.01-403.14 |
References Cited
[Referenced By]
U.S. Patent Documents
Primary Examiner: Noguerola; Alex
Attorney, Agent or Firm: Greenblum & Bernstein,
P.L.C.
Claims
The invention claimed is:
1. A biosensor comprising: an electrically insulating base plate; a
plurality of lead terminals formed on the base plate; a plurality
of lead wires connected to the lead terminals, respectively; an
electrode system including two working electrodes and one reference
electrode connected to the lead wires, respectively; an insulating
layer that insulates the electrodes; an enzyme reaction layer
formed on the insulating layer and the electrodes; a spacer formed
on the enzyme reaction layer so as to ensure a sufficient space to
receive a sample; and a cover formed on the spacer, wherein the
spacer has a sample introduction port opened at one side of the
spacer, and the cover has at least one slit that vents air existing
in a sample receiving space defined by the spacer and the cover,
the slit extending to above the electrodes from one end of the
cover.
2. The biosensor as set forth in claim 1, further including a
curved groove formed at one end of the cover toward the electrodes,
the slit being formed from a certain position of the curved groove
with a predetermined length.
3. The biosensor as set forth in claim 2, wherein the slit is a
plurality of slits spaced at a predetermined interval along a
length direction.
4. The biosensor as set forth in claim 2, wherein the respective
working electrodes have the same electrical resistance and area,
and the reference electrode is spaced at the same distance from the
respective working electrodes.
5. The biosensor as set forth in claim 4, wherein the reference
electrode has an area more than 1.5 times larger than the working
electrodes.
6. The biosensor as set forth in claim 1, wherein the slit is a
plurality of slits spaced at a predetermined interval along a
length direction.
7. The biosensor as set forth in claim 1, wherein the respective
working electrodes have the same electrical resistance and area,
and the reference electrode is spaced at the same distance from the
respective working electrodes.
8. The biosensor as set forth in claim 7, wherein the reference
electrode has an area more than 1.5 times larger than the working
electrodes.
9. The biosensor as set forth in claim 1, wherein when the sample
receiving space is placed in a vertical direction, the reference
electrode has an H-shape, and the respective working electrodes are
arranged at the upper and lower portions of a horizontal axis of
the reference electrode.
10. The biosensor as set forth in claim 9, wherein the respective
working electrodes have the same electrical resistance and area,
and the reference electrode is spaced at the same distance from the
respective working electrodes.
11. The biosensor as set forth in claim 1, wherein when the sample
receiving space is placed in a vertical direction, the reference
electrode has an E-shape, and the respective working electrodes are
arranged at the upper and lower portions of a horizontal axis,
which is formed at the center of the reference electrode.
12. The biosensor as set forth in claim 11, wherein the respective
working electrodes have the same electrical resistance and area,
and the reference electrode is spaced at the same distance from the
respective working electrodes.
Description
This application claims priority under 35 U.S.C. .sctn. 371 from
PCT/KR02/01854, filed on Oct. 4, 2002 and under 35 U.S.C. .sctn.
119(a d) from Korean patent application KR 2002/27971, filed on May
20, 2002.
TECHNICAL FIELD
The present invention relates to a sensor, and more specifically to
a biosensor for quantifying a specific substance contained in a
biological sample.
BACKGROUND ART
Generally, a biosensor comprises an electrically insulating base
plate, an electrode system including a plurality of electrodes and
formed on the electrically insulating base plate using a screen
printing method, and an enzyme reaction layer including a
hydrophilic polymer, oxidoreductase and an electron acceptor and
formed on the electrode system. When a sample liquid containing a
substrate is dropped on the enzyme reaction layer of the biosensor,
the enzyme reaction layer is dissolved to allow the substrate and
enzyme to react with each other. At a result, the substrate is
oxidized, and then the electron acceptor is reduced. After such an
enzyme reaction finishes, the concentration of the substrate in the
sample liquid is determined from an oxidation current obtained by
electrochemically oxidizing the reduced electron acceptor.
As a biosensor for quantifying a specific substance contained in a
biological sample using an electrochemical manner, a glucose sensor
is known. FIGS. 1 and 2 show a structure of the glucose sensor.
FIG. 1 is an exploded perspective view of a conventional biosensor
in which a reaction layer is omitted. FIG. 2 is a longitudinal
sectional view of the biosensor shown in FIG. 1.
Referring to FIG. 1, silver paste is screen-printed on an
electrically insulating base plate 1 to form leads 2 and 3 on a
base plate 1. Conductive carbon paste containing a resin binder is
then printed on the base plate 1 to form an operating electrode 4
on the base plate 1. The operating electrode 4 is contacted with
the lead 2. Electrically insulating paste is then printed on the
base plate 1 to form an insulating layer 6. The insulating layer 6
covers all portions except the operating electrode 4 so that the
exposed area of the operating electrode 4 is maintained to be
constant. Conductive carbon paste containing the resin binder is
printed on the base plate 1 to come into contact with the lead 3
and thus to form a ring-shaped counter electrode 5. Subsequently,
on or near an electrode system including the operating electrode
and the counter electrode, a reaction layer is formed.
The electrically insulating base plate 1 having the reaction layer
and a cover 9 having an air hole 11 are bonded to each other via a
spacer 10, along dashed dot lines marked in FIG. 1, to manufacture
a biosensor. A slit 13 is formed at the spacer 10 to provide a
sample supplying path between the base plate and the cover.
Referring to a longitudinal sectional view of the biosensor having
the above-mentioned structure, a hydrophilic polymer layer 7 is
disposed at the electrically insulating base plate 1 having the
electrode system, and a reaction layer 8 including enzymes and
electron acceptors and a lecithin layer 8a are disposed on the
hydrophilic polymer layer 7 in this order.
When a biological sample is contacted with an introduction port 12
of the biosensor having the above-mentioned structure, the
biological sample fills the slit 13 acting as a sample receiving
space, and at the same time air in the sample receiving space is
vented through an air hole 11 formed at the cover 9.
However, since the air hole 11 is formed at the upper part of the
biosensor, the biosensor is disadvantageous in terms of its
handling due to measurement errors caused by frequent contact with
the air hole 11 when using the biosensor. Considering the fact that
the reaction progresses immediately after the sample comes into
contact with the reaction layer, it is important to rapidly absorb
the sample irrespective of viscosity of the sample. However, in the
biosensor having the above-mentioned structure, since the air hole
11 for venting air is arranged at the rear side of a sample
introduction passage, rapid absorption of the sample is limited.
Such limited absorption of the sample causes measurement errors in
biosensors that initiate the measurement after checking whether or
not the sample is completely introduced.
DISCLOSURE OF THE INVENTION
Therefore, the present invention has been made in view of the above
problems, and it is an object of the present invention to provide a
biosensor capable of rapidly inducing absorption of a biological
sample, thereby minimizing measurement errors caused by the
biosensor.
It is another object of the present invention to provide a
biosensor which can provide simplicity and convenience in handling
the biosensor and accurately measure a reactive substance contained
in a sample using two working electrodes and one reference
electrode.
To achieve the above objects, there is provided a biosensor
comprising: an electrically insulating base plate 20; a plurality
of lead terminals 32 formed on the base plate 20; a plurality of
lead wires 31 connected to the lead terminals 32, respectively; an
electrode system 40 including two working electrodes 42 and 43 and
one reference electrode 41 connected to the lead wires 31,
respectively; an insulating layer 50 for insulating the electrodes
41, 42 and 43; an enzyme reaction layer 80 formed on the insulating
layer 50 and the electrodes 41, 42 and 43; a spacer 60 formed on
the enzyme reaction layer 80 so as to ensure a sufficient space for
receiving a sample; and a cover 70 formed on the spacer 60,
wherein the spacer 60 has a sample introduction port 61 formed at
one side of the spacer 60, and the cover 70 has at least one slit
71 for venting air existing in a sample receiving space 62 defined
by the spacer 60 and the cover 70, the slit 71 extending to above
the electrodes 41, 42 and 43 from one end of the cover 70.
In the biosensor according to the embodiment of the present
invention, since the sample receiving space 62 is opened through
the slit 71, a biological sample is rapidly introduced into the
biosensor by virtue of maximized capillary effect.
In accordance with one aspect of the present invention, there is
provided a biosensor further including a curved groove 72 formed at
one end of the cover 70 toward the electrodes 41, 42 and 43, the
slit 71 being formed from a certain position of the curved groove
72 with a predetermined length. The biosensor having the
above-mentioned structure can collect a quantity of biological
sample in the curved groove 72.
In accordance with the embodiment of the biosensor according to the
present invention, the respective working electrodes 42 and 43 have
the same electrical resistance and area, and the reference
electrode 41 is spaced at the same distance from the respective
working electrodes 42 and 43 and has an area above 1.5 times larger
than the working electrodes 42 and 43.
BRIEF DESCRIPTION OF THE DRAWINGS
The above and other objects, features and other advantages of the
present invention will be more clearly understood from the
following detailed description taken in conjunction with the
accompanying drawing, in which:
FIG. 1 is an exploded perspective view of a conventional biosensor
in which a reaction layer is omitted;
FIG. 2 is a longitudinal sectional view of the biosensor shown in
FIG. 1;
FIGS. 3a and 3b are a top view and a back view of a biosensor
according to an embodiment of the present invention,
respectively;
FIG. 4 is an exploded perspective view of a biosensor according to
an embodiment of the present invention;
FIG. 5 is a cross-sectional view of a biosensor according to an
embodiment of the present invention;
FIG. 6 is a front view of the biosensor when viewed from a
direction A (shown by an arrow) in FIG. 5; and
FIGS. 7a to 7e are diagrams showing various electrode arrangements
and positions of the slit for venting air in a biosensor according
to an embodiment of the present invention.
BEST MODE FOR CARRYING OUT THE INVENTION
Hereinafter, the present invention will be explained in more detail
through preferred embodiments, with reference to the accompanying
drawings in such a manner that it may easily be carried out by a
person having ordinary skill in the art.
FIGS. 3a and 3b are a top view and a back view of a biosensor
according to an embodiment of the present invention,
respectively.
Referring to FIG. 3a, a plurality of lead terminals 31
corresponding to the number of electrodes are formed at one end of
an electrically insulating base plate 20. As shown in FIG. 3b, the
lead terminals 31 are connected to electrodes 41, 42 and 43,
respectively, formed at the other end of the electrically
insulating base plate 20 through respective lead wires 32. As shown
in FIG. 3a, a slit 71 is formed at a cover 70 of the biosensor S
according to the embodiment of the present invention, and extends
from a curved groove 72 formed at one end of the cover 70 toward
the electrodes 41, 42 and 43 to at least above the electrodes 41,
42 and 43. The slit 71, which will be discussed below, acts as an
air-vent when a biological sample is introduced by the capillary
phenomenon.
The electrically insulating base plate 20 may be made of a
non-conductive material such as polyethylene terephthalate,
polyvinyl chloride resin, polycarbonate resin, etc. A lead section
30 including the lead wires 32 and the lead terminals 31 may be
formed in accordance with a common method such as screen printing.
In the embodiment of the present invention, the lead section 30 was
formed by screen-printing a silver ink or a mixed ink of silver and
silver chloride on the base plate 20.
The structure of the biosensor S will now be explained in more
detail with reference to the exploded perspective view and
cross-sectional view of FIGS. 3a and 3b.
FIG. 4 is an exploded perspective view of the biosensor S according
to the embodiment of the present invention, and FIG. 5 is a
cross-sectional view of the biosensor according to the embodiment
of the present invention.
Referring to FIG. 4, the biosensor S according to the embodiment of
the present invention comprises the electrically insulating base
plate 20, the base plate 20 having three lead terminals 32 formed
on one end of the base plate 20, the lead terminals 32 being
connected to three electrodes 41, 42 and 43, respectively, through
the lead wires 31. In the electrodes 41, 42 and 43, a reference
numeral 41 denotes a reference electrode, and reference numerals 42
and 43 denote working electrodes. These electrodes act to measure
the amount of current generated during oxidation and reduction of
an electron acceptor included in an enzyme reaction layer 80, which
will be discussed below. The reference electrode 41 is arranged
between the respective working electrodes 42 and 43. This electrode
arrangement makes it possible to measure the amounts of current in
the reference electrode 41 and the respective working electrodes 42
and 43. That is, the biosensor according to the embodiment of the
present invention measures the amounts of current between the first
working electrode 43 and the reference electrode 41, and the second
working electrode 42 and the reference electrode 41, compares the
measured values and determines errors generated in the manufacture
of the biosensor and the reaction with a substrate, thereby
quantitatively obtaining the concentration of the substrate
contained in the biological sample with an increased accuracy.
In accordance with the embodiment of the biosensor according to the
present invention, in order to measure the amounts of current in
the reference electrode 41 and the respective working electrodes 42
and 43 under the same electrochemical conditions, the respective
working electrodes 42 and 43 must have the same electrical
resistance and area, and the reference electrode 41 must be spaced
at the same distance from the respective working electrodes 42 and
43. In addition, the area of the reference electrode (41) is
preferably more than 1.5 times larger than that of the working
electrodes 42 and 43. Since the amounts of current generated in the
reference electrode 41 and the respective working electrodes 42 and
43 is proportional to the reactive area of the electrodes, the
relatively large area of the reference electrode 41 can reduce
measurement errors between the reference electrode 41 and the
respective working electrodes 42 and 43. The reference electrode 41
and the working electrodes 42 and 43 are collectively referred to
as "an electrode system 40". The electrode system 40 can be formed
by a screen printing method using a conductive carbon ink.
In order to insulate the electrodes 41, 42 and 43, an insulating
material is partially coated on the electrodes 41, 42 and 43 except
the upper portions of the electrodes 41, 42 and 43 to form an
insulating layer 50, as shown in FIG. 5. As the insulating
material, a non-conductive ink for screen printing or an ink for
insulation can be used. The enzyme reaction layer 80 is formed on
both the exposed portions of the electrodes 41, 42 and 43 and the
insulating layer 50. The enzyme reaction layer 80 includes an
enzyme reactive with the introduced biological sample, and an
electron acceptor.
The enzyme reaction layer 80 must include an enzyme reactive with a
substrate to be detected. That is, the enzyme reaction layer 80 can
include different enzymes depending on the application of the
biosensor. Examples of the enzymes and substrates are shown in
Table 1 below. As shown in Table 1, when the biosensor according to
the embodiment of the present invention is a glucose sensor, the
enzyme reaction layer 80 includes glucose oxidase. When a blood
sample as the biological sample is introduced into the enzyme
reaction layer 80 of the sensor, glucose in blood is oxidized by
glucose oxidase, after which the glucose oxidase is reduced.
Herein, the electron acceptor included in the enzyme reaction layer
80 oxidizes the glucose oxidase and then itself is reduced. The
reduced electron acceptor loses its electrons on the surface of the
electrode, to which a constant voltage is applied, and then is
electrochemically reoxidized. Since the concentration of glucose in
the blood sample is proportional to the amount of current generated
when the electron acceptor is oxidized, the concentration of
glucose in the blood sample can be measured by measuring the amount
of current through the lead terminals 32.
TABLE-US-00001 TABLE 1 Substrate Enzymes Glucose Glucose oxidase
Cholesterol Cholesterol esterase Cholesterol oxidase Peroxidase
Creatinine Creatininase Creatinase Sarcosine oxidase Lactate
Lactate oxidase
On the other hand, in accordance with the biosensor S according to
the embodiment of the present invention, a spacer 60 having a
sample introduction port 61 for forming a sample receiving space is
formed on the enzyme reaction layer 80, and is sandwiched between
the base plate 20 and the cover 70. In order to form the sample
receiving space 62 between the cover 70 and the enzyme reaction
layer 80 when the cover 70 and the spacer 60 are bonded to each
other, the spacer 60 must be higher than the enzyme reaction layer
80 formed on the base plate 20. The spacer 60 can be made of resin.
In the embodiment of the present invention, a double-sided tape
made of resin was used as the spacer 60.
In accordance with the biosensor S according to the embodiment of
the present invention, the cover 70 is bonded to the spacer 60. At
this time, in order to vent air existing in the sample receiving
space 62 between the spacer 60 and the cover 70, the slit 71 is
formed at the cover 70. For stable introduction of the biological
sample into above the electrode 42, the slit 71 extends to at least
above the electrodes 41, 42 and 43 with a predetermined length.
In the biosensor S as shown in FIG. 5, the spacer 60 is bonded to
the upper side of the insulating layer 50. However, the spacer 60
can be directly bonded to the base plate 20 instead of the
insulating layer 50.
FIG. 6 is a front view of the biosensor when viewed from a
direction A in FIG. 5.
Referring to FIG. 6, the base plate 20 is placed at the bottom of
the biosensor S according to the embodiment of the present
invention, the insulating layer 50 insulates the electrodes 41, 42
and 43 formed on the base plate 20, and the enzyme reaction layer
80 is formed on the insulating layer 50. Since the spacer 60 is
higher than the enzyme reaction layer 80 and is placed around the
electrodes 41, 42 and 43, the sample receiving space 62 is formed
between the cover 70 and the enzyme reaction layer 80. Since the
slit 71 formed at the cover 70 acts as an air-vent, the biological
sample is introduced into the sample receiving space 62 by virtue
of capillary phenomenon.
FIGS. 7a to 7e are diagrams showing various electrode arrangements
and positions of the slit 71 for venting air in the biosensor S
according to the embodiment of the present invention.
In the biosensor S according to the embodiment of the present
invention, the electrode system 40 including the reference
electrode 41 and the working electrodes 42 and 43 can be formed as
shown in FIG. 7a. That is, the reference electrode 41 has an
E-shape, and the respective working electrodes 42 and 43 can be
arranged at the upper and lower portions of a horizontal axis B,
which is formed at the center of the reference electrode 41. In
this case, the respective working electrodes 42 and 43 must also
have the same electrical resistance and area, and the reference
electrode 41 must be spaced at the same distance from the
respective working electrodes 42 and 43.
In accordance with another embodiment of the present invention, the
electrode system 40 can be formed as shown in FIG. 7b. That is,
when the sample receiving space 62 is placed in a vertical
direction, the reference electrode 41 has an H-shape, and the
respective working electrodes 42 and 43 can be arranged at the
upper and lower portions of a horizontal axis of the reference
electrode 41. Also, the respective working electrodes 42 and 43
must have the same electrical resistance and area, and the
reference electrode 41 must be spaced at the same distance from the
respective working electrodes 42 and 43 and have an area above 1.5
times larger than the working electrodes 42 and 43.
In the electrode systems shown in FIGS. 7a and 7b, the slit 71
formed at the cover 70 extends to above the reference electrode 41
and the working electrodes 42 and 43.
FIGS. 7c to 7e are diagrams showing various positions of the slit
71 formed at the cover 70. For example, FIG. 7c shows the slit 71
formed along a length direction of the cover 70, FIG. 7d shows a
plurality of slits 71 formed at the center of the cover 70, each
slit being spaced at a predetermined interval, and FIG. 7e shows
the slit 71 formed along the right side of the cover 70. All slits
71 shown in FIGS. 7c to 7e extend to above the electrodes 41, 42
and 43 arranged on the base plate 20, thereby stably introducing
the biological sample into the upper portion of the electrodes 41,
42 and 43.
Operation of a glucose sensor as an example of the biosensor
according to the embodiment of the present invention will be
explained below.
Referring to FIG. 3a, after the curved groove 72 of the glucose
sensor is contacted with a blood sample, the blood sample is
introduced into the sample introduction port 61 by virtue of
capillary phenomenon and simultaneously air existing in the sample
receiving space 62 is vented through the slit 71 formed at the
cover 70. Subsequently, the blood sample is introduced into the
first working electrode 43, the reference electrode 41 and the
second working electrode 42 through the sample receiving space 62,
and the blood sample is then impregnated into the enzyme reaction
layer 80. Glucose contained in the blood sample enzymatically
reacts with GOD, so that it is oxidized and simultaneously the GOD
is reduced. The reduced GOD reacts with the electron acceptor and
is reoxidized, after which the reoxidized GOD reacts with other
glucose not yet oxidized. The reduced electron acceptor migrates to
the surface of the electrode, to which a voltage is applied, and
then loses electrons at the surface to be electrochemically
reoxidized. Thereafter, the electron acceptor continuously takes
part in the above reaction. The current generated during the
oxidation of the electron acceptor is proportional to the
concentration of glucose in the blood sample. Accordingly, the
concentration of glucose in the blood sample can be more accurately
and quantitatively obtained by measuring the amounts of current
flowing in the first working electrode 43 and the reference
electrode 41, and the second working electrode 42 and the reference
electrode 41, respectively, and averaging the measured values.
To facilitate introduction of sample into the biosensor, the
biosensor according to the present invention has the slit 71 formed
at the cover, instead of an air hole. Accordingly, the biosensor
according to the present invention can provide simplicity and
convenience in handling the biosensor. In addition, since the slit
71 extends from one end of the cover 70 to above the electrodes,
the sample can be rapidly introduced into the electrodes.
INDUSTRIAL APPLICABILITY
As described above, since the biosensor according to the present
invention has an air-venting slit extending from one end of a cover
to above the electrodes, a biological sample can be rapidly
introduced. In addition, measurement errors caused by the biosensor
can be minimized by measuring the amounts of current flowing in the
two electrodes and one reference electrode, and averaging the
measured values. Furthermore, since the biosensor according to the
present invention does not include a separate air hole for
facilitate introduction of a sample into the biosensor, it can
provide simplicity and convenience in handling the biosensor.
While the present invention has been described with regard to
preferred embodiments thereof, the description is for illustrative
purposes only and is not to be construed as limiting the scope of
the invention. Various modifications and changes may be made by
those skilled in the art without departing from the true scope of
the invention as defined by the appended claims.
* * * * *